Investigation of ionic channel and its * cance in *

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1) Electro-physiological effects of endothelin-1 on human bronchial smooth muscle cells : We investigated the effects of endothelin-1 and histamine on electro-physiological effects of endothelin-1 on human bronchial smooth muscle cells, by means of the patch-clamp techniques and Ca^<2+> measurements using fura-2 AM, Both endothelin-1 and histamine first increased [Ca^<2+>]_1 by the release of Ca^<2+> from store sites, then fol1owed by the sustained Ca^<2+> influx. The [Ca^<2+>]_1 increase induced by endothelin-1 was mediated by both ET_A and ET_B receptors. The activation of Ca^<2+>-permeable nonselective cation channels (I_<NS>) was involved in the sustained rise in [Ca^<2+>]_1, where ET_A and ET_B receptors were involved. The EC_<50> value for endothelin-1 to activate I_<N,S-> was approximately 10 nM.In addition, presence of both ET_A and ET_B mRNA was confirmed by RT-PCR in these cells and immunostaining of ET receptors in human airway preparations.2) K^+ channels in human bronchial smooth muscle cells : We examined the properties of K^+ channels, by using the patch clamp techniques and the RT-PCR analysis of mRNA and antisense oligonucleotide methods. In these cells, we found the voltage-dependent K^+ channels (K_v) and the inwardly rectifying K^+ channels (Kir), which contribute to forming the resting membrane potential. The Kir channels was blocked by low doses of extracellular Ba^<2+> ions with a IC_<50> of approximately - 1.8μM.RT-PCR analysis of mRNA showed transcripts for Kir2.1. but not Kir1.1, 2.2, and 2.3. Treatment of cells with antisense oligonucleotides targeted to Kir2.1 resulted in a significant decrease in current density of the Kir current. These results demonstrate that the inward rectifier K^+ current (Kir) is present in human bronchial smooth muscle cells. and the Kir2.1 gene encodes the Kir channel protein in these cells.